Oligofructose (OF), comprised of fructose oligomers with a terminal glucose unit, is a family of oligosaccharides derived from the hydrolysis of inulin. Consumption of OF in animals and humans increases colonic bifidobacteria levels. The present study evaluates the safety of OF in both a 13 week rat feeding study and using in vitro mutagenicity tests. Fecal bifidobacteria levels were also determined by in situ hybridization to assess a biological function of OF. Rats received either a control diet or diets containing one of four doses of OF. Total, HDL, and LDL-cholesterol levels were significantly lower at several time points during the study in groups receiving OF compared to controls with the largest effects occurring in the high dose male animals. Weight gain in the male high dose group was significantly lower at early time points compared to controls but not significantly different at the end of study. As expected, cecal weights increased in a dose-related manner and fecal bifidobacteria levels also demonstrated a dose-related increase. There were no consistent differences in gross pathology or histopathology related to dietary OF. OF did not induce a positive response in the Ames test or chromosomal aberration test with CHO cells. These results demonstrate no adverse effects of OF.